Analysis and development of L-threonine-overproducing escherichia coli based on functional genomics and metabolic engineering = 기능 유전체학 및 대사공학을 이용한 쓰레오닌 과생산 대장균의 분석 및 개발

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We compared the transcriptome, proteome, and nucleotide sequences between wild-type E coli W3110 and the L-threonine-overproducing mutant E. coli TF5015. DNA macroarrays were used to measure mRNA levels for the all genes of E. coli and two-dimensional gel electrophoresis was employed to compare protein levels. It was observed that only 54 in 4290 genes (1.3%) exhibited differential expression profiles. Typically, genes such as aceA, aceB, icdA, gltA, glnA, leu operon, proA, thrA, thrC, and yigJ, which are involved in glyoxylate shunt, TCA cycle, and amino acids biosynthesis (L-glutamine, L-leucine, proline, and L-threonine), were significantly up-regulated, while the genes dadAX, hdeA, hdeB, ompF, oppA, oppB, oppF, yfiD, and many ribosomal protein genes were down-regulated in TF5015 compared to W3110. The differential expression such as up-regulation of thr operon, repression of asd, dapB, and metE, and expression of yigJ would result in an accumulation of L-threonine in TF5015. Furthermore, two significant mutations, thrA345 and ilvA97, which are essential for overproduction of L-threonine, were identified in TF5015 through the sequence analysis. Up-regulation of aceBA and down-regulation of b1795, hdeAB, oppA, and yfiD seem to be linked with a low accumulation of acetate in TF5015. Such comprehensive analyses provide information to understand the regulatory mechanism of L-threonine production and the physiological consequences in the mutant stain. Based on the functional genomic analysis and biochemical information, we reconstructed the metabolic map for L-threonine in TF5015, and determined the target genes to be modified for metabolic overproduction of L-threonine in TF5015. Of the selected target genes, introduction of an extra copy of the thr operon or the asd, ppc, and aspC into TF5015 resulted in an increase of L-threonine production by 12%, 6%, 5%, and 4.5%, respectively. These results support our approach and suggest that it can be effectively used fo...
Advisors
Kim, Hak-Sungresearcher김학성researcher
Description
한국과학기술원 : 생물과학과,
Publisher
한국과학기술원
Issue Date
2003
Identifier
180988/325007 / 000985291
Language
eng
Description

학위논문(박사) - 한국과학기술원 : 생물과학과, 2003.2, [ vii, 93 p. ]

Keywords

Functional genomics; Threonine; Metabolic engineering; 대사공학; 기능 유전체학; 쓰레오닌

URI
http://hdl.handle.net/10203/27541
Link
http://library.kaist.ac.kr/search/detail/view.do?bibCtrlNo=180988&flag=dissertation
Appears in Collection
BS-Theses_Ph.D.(박사논문)
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